Project Summary We propose the existence of a ?Microtubule Integrity Response? (MIR), akin to the DNA damage and unfolded protein responses. The MIR network, in our hypothesis, comprises a system of sensor molecules that detect perturbation of microtubules and/or soluble tubulin by environmental insults and normal physiological inputs. In response, they control downstream signaling and gene expression. We will combine biochemical and genetic approaches to identify MIR sensors, determine how they control signaling and gene expression, and explore the consequences for microtubule physiology and pharmacology. We will identify a hypothetical microtubule- bound kinase whose phosphorylation of MAPs and tip-tracking proteins depends on intact microtubules, and test its role in homeostatic regulation of microtubules. We will determine how multiple Jnk-dependent phosphosites in the nucleus are increased following microtubule stabilization, and the consequences for cell physiology. We identified a robust MIR mRNA signature in which mRNAs for multiple ??? and ?-tubulins are counter-regulated by MT stabilizing vs destabilizing drugs across multiple cell types. We will use this signature in bioinformatics searches to discover novel microtubule physiology, e.g metabolic regulation of microtubules. We will develop imaging biosensors to measures soluble Tb and its spatiotemporal regulation in single cells, and homeostatic response to pertrubation. Finally, we will test the hypothesis that microtubules serve as sensors for mechanical cues such as cell shape, substrate stiffness and cytoplasmic crowding, and control cell responses to these cues via MIR signaling. Elucidating the molecular- and cell biology of the MIR will open new directions in fundamental cytoskeleton research, help us understand the therapeutic and toxic actions of microtubule targeting drugs using in cancer and inflammation, and reveal novel human physiology.